Our overall hypothesis is that a defense-in-depth strategy, according to which invading HIV meets not only strong resistance at mucosal frontlines but also at various deeper levels, will greatly improve AIDS vaccine efficacy. Although ~90% of all HIV transmissions occur mucosally, only few studies have focused on the initial events as HIV enters mucosal fluids and crosses epithelial barriers, and on the role of mucosal IgA, which is mostly dimeric (dIgA). We gave the first direct evidence that dIgA is protective when we passively immunized rhesus monkeys (RMs) intrarectally (i.r.) with the monoclonal antibody (mAb) HGN194 as dIgA1, dIgA2, or IgG1 forms; all neutralized the clade C SHIV (SHIV-C) challenge virus equally well in vitro. Given i.r., dIgA2 protected 17% and dIgA1 83% of the RMs against i.r. SHIV-C challenge, respectively; better protection was linked to virion capture and inhibition of transcytosis of cell-free virus. Next, we combined i.v. HGN194 IgG1 with i.r. dIgA2 and compared this combination with i.v. IgG1 nmAb given alone. The latter protected none of the RMs, whereas 100% of the RMs given the IgG1+dIgA2 combination were protected. This strong synergy prompted us to study the interaction between virions, the various Abs in mucosal fluids, and mucus (Project 1). The association of better protection with virion capture, over and above neutralization, led us to postulate that non-neutralizing dIgAs with potent virion-binding ability could prevent mucosal transmission (Project 2). Recently, Darrell Irvine targeted polypeptide antigens and molecular adjuvants to lymph nodes (LNs) by conjugating both to amphiphilic (amph) albumin-binding lipid tails, resulting in efficient albumin-mediated LN uptake. Such amph-vaccines increased T-cell responses 30-fold and humoral responses 20-fold and improved the therapeutic efficacy of cancer vaccines in mice. This P01 seeks to test the hypothesis that immunogens targeted to draining LNs of mucosal virus entry sites will generate strong mucosal IgA defenses as well as IgG and cellular immune responses (Project 3). LN targeting will be followed over time by positron emission tomography (PET) in vaccinated RMs in the Research Imaging Institute (Core D, Drs. Fox & Goins). Cores B (Virology/Immunology; Dr. Sholukh and Lanzavecchia) and C (Primate Studies; Drs. Lakhashe & Gupta) will support all Projects. We seek to test our hypotheses with these Specific Aims: 1. to gain a greater understanding of the initial steps as virus enters the mucosal lumen, interacts with mucus, mucosal Abs, and the cells at and just beyond the epithelial barrier (Project 1; Dr. Hope), 2. to passively administer mAbs with epitope specificities linked to protection and inducible by currently available immunogens, including non-neutralizing epitopes, to examine the interaction of systemically administered IgGs with i.r. administered dIgAs (Project 2; Dr. Ruprecht), and 3. to target vaccine-induced immunity to LNs and mucosal compartments (Project 3; Dr. Irvine).